A NON-DESTRUCTIVE METHOD OF SAMPLING LIVE CORAL TISSUE, SINGLE POLYP-SAMPLE PREPARATION AND PROTEIN QUANTIFICATION FOR ASSESSMENT OF CORAL HEAT SHOCK PROTEINS

摘要: Sessile marine invertebrates are routinely exposed to fluctuations in their environment such as temperature, irradiance, terrestrial run-off and water-flow. Many of these fluctuations can affect a coral reef at a range of scales from molecular to an entire reef system with resulting effects on growth and energetics, reproductive efficiency, protein damage and the loss of pigment or expulsion of the symbiotic dinoflagellate Symbiodinium. The induction and regulation of heat shock proteins (hsps), is a significant defense mechanism that can preserve metabolic function and foster recovery from short-term stress events. Adaptation from thermal histories can help corals in subsequent thermal insults. Current coral sampling methods often employ an invasive approach in the collection of coral tissue, cleaving off coral fragments (~ 100 cm2) from live coral colonies that may already be in a stressed or poor condition. Subsequent protein preparative methods have to further purify the sample to account for contaminants, eg, CaC03and coral mucus. In the present study, three methods were developed specifically to;(i) carryout single polyp time-series sampling on live coral colonies (< 150 cm2)(n= 6), without compromising the colonies or parallel studies on the photobiology of the colonies,(ii) develop a low volume (50-100 pi of coral tissue) protein recovery method for single coral polyps, and (iii) develop a low volume, high resolution protein quantification method. The preliminary testing of five separate protein preparation methods resulted in a range of total protein yields from 47 to 77 (ug) per